The liver is the key organ in the body that regulates vitamin A homeostasis. It is the tissue where approximately 75% of newly absorbed dietary vitamin A is taken up; the site where approximately 80-90% of the vitamin A present in the body of a healthy well nourished individual is stored; and the site from which stored vitamin A is mobilized to meet the needs of extrahepatic tissues. Newly absorbed vitamin A arriving as retinyl ester in chylomicron remnants is taken up by hepatocytes and must be hydrolyzed to retinol before it is transferred to hepatic stellate cells (HSCs) for storage. Approximately 80% of hepatic vitamin A exists as retinyl ester in the lipid droplets of HSCs. HSC retinyl ester stores must be hydrolyzed to retinol so that vitamin A can be mobilized into the circulation bound to retinol-binding protein (RBP). Although there has been longstanding research interest focused on identifying the hepatic enzyme(s) responsible for catalyzing retinyl ester hydrolysis, the molecular identity(ies) of this enzyme(s) remains elusive. Our overall goal is to identify at the molecular level the enzyme(s) responsible in vivo for hydrolyzing chylomicron remnant retinyl ester in hepatocytes and lipid droplet retinyl esters in HSCs. The molecular identify of the enzyme which catalyzes retinyl ester hydrolysis in HSC lipid droplets and the factors that modulate hepatic retinol mobilization will be explored in Specific Aim 1. The molecular identity of the enzyme which catalyzes hydrolysis of newly absorbed chylomicron retinyl ester will be investigated in Specific Aim 2. Clinical Relevance. Over the past decade, a large literature reporting relationships between high blood retinol and RBP levels and metabolic disease, especially obesity, insulin resistance, liver disease, and cardiovascular disease has accumulated. Most of this literature consists of retrospective and prospective studies of human cohorts, but animal model and cell culture studies also point to these same relationships. Yet, the factors which control retinol and RBP release from the liver, and consequently blood levels, are not understood at the molecular level. We are proposing to investigate an enzymatic process, hepatic retinyl ester hydrolysis, that is essential to the regulation of blood retinol and RBP levels. Global Health Relevance. Vitamin A-deficiency remains a major global health problem. The World Health Organization estimates that millions of infants and young children worldwide are at risk of vitamin A-deficiency. We are proposing to study a process, the hydrolysis of hepatic vitamin A esters, that is central for allowing the body to maintain normal blood and tissue vitamin A levels, thus preventing vitamin A-deficiency. Our studies will advance understanding of an enzymatic process that is critical for allowing vitamin A to be accumulated in the liver and to be mobilized from the liver in order to keep cells and tissues healthy.